Bacterial resistance to antibiotics has been a great problem for many years. The degree of resistance and the speed with which resistance develops varies with different organisms and different drugs. Enzymes called β-lactamases are produced by bacteria and are one mechanism in which bacteria develop antimicrobial resistance. Gram-negative bacteria producing enzymes called ESBLs because of their wide substrate range are of a particular concern in nosocomial infections. In many countries people still use traditional medicine derived from plants as an alternative to the Western medicine due to increased cost of Western medicine and microbial resistance of antibiotic treatments. Biologically active compounds isolated from plants species are used in herbal medicine. Because of the high prevalence of the ESBLs and their increasing resistance to the antibiotics, this research study was done to test the antimicrobial activities of selected medicinal plants of the Eastern Cape; G. incanum, D. angustifolia and E. autumnalis which were traditionally used to treat various infections. The in vitro antimicrobial activity of three different extracts (acetone, methanol & distilled water) and the traditional preparations of the three plants were tested against the selected strains of ESBL-producing bacteria, non β-lactamase producers and the different fungal species. The extracts were screened against 26 Gram-positive bacterial strains, 53 Gram-negative bacterial strains and 15 fungal strains. The Gram-positive bacteria included strains from S. aureus, B. cereus and E. faecalis. The Gram-negative bacteria included strains from E. ii coli, E. cloacae, K. pneumoniae, P. aeruginosa and Acinetobacter spp. The fungal strains included 9 strains of Candida albicans and a single strain of each of the following opportunistic fungi, Mucor sp, Geotrichium sp, Penicillium sp, Fusarium sp and Rhizopus sp. The agar dilution assay was used for the antimicrobial screening of the plants extracts and for the determination of the MICs. The Ames test was performed for the determination of probable carcinogenicity of the extracts of G. incanum and D. angustifolia. The distilled water extracts followed by acetone extracts of the plants revealed the highest antimicrobial activity against the different microbial strains. The extracts of G. incanum followed by the extracts of D. angustifolia inhibited the highest number of microbial strains. The extracts of E. autumnalis did not show any antimicrobial activity against all the pathogens in this study. More of the Gram-positive bacteria were inhibited by the plant extracts. The lowest MIC was obtained with Gram-positive bacteria. The bacterial strains of E. faecalis and P. aeruginosa were not inhibited by any of the plants extracts in the agar dilution assay yet Acinetobacter species which are MDR were inhibited by the distilled water and methanol extracts of G. incanum. A single strain of Mucor sp was the only spore forming fungi that was inhibited by the distilled water extracts of G. incanum. None of the plants extracts showed any mutagenic effects on the TA100 S. typhimurium strains incorporated on the Ames test. Apart from revealing of new antimicrobial agents that may be used against resistant organisms, the proper use of antimicrobial agents should be recommended. The study has highlighted a need for further investigations on the properties of the medicinal plants used in this study.

Rapidly emerging strains of bacteria resistant to most advanced antibiotics have become issues of very important public health concern. Research currently directed towards marine actinomycetes presents a vast potential for new compounds that could be able to safely and effectively target resistant species. In this regard, ten putative Streptomyces strains isolated from the Nahoon beach were selected and assessed for antibiotic production and activity against a wide range of bacteria including reference strains, environmental strain and clinical isolates. The ethyl acetate extracts of the putative Streptomyces isolates showed activities against at least 6 and up to 26 of the 32 test bacteria. Inhibition zones were found to range between 9-32 mm diameters at a concentration of 10 mg/ml. The minimum inhibitory concentrations (MICs) of the crude extracts ranged from 0.039 - 10 mg/ml and the least minimum bactericidal concentration (MBC) demonstrated was 0.625 mg/ml against a reference strain Staphylococcus aureus ATCC 6538. Time kill kinetics of all extracts revealed bacteristatic and bactericidal activities. Average Log reductions in viable cell counts for all the extracts ranged from 0.86 Log10 and 3.99 Log10 cfu/ml after 3 h interaction and 0.01 Log10 and 4.86 Log10 after 6 h interaction at MIC, 2 × MIC, 3 × MIC and 4 × MIC concentrations. Most of the extracts were speedily bactericidal at 3 × MIC and 4 × MIC resulting in over 50 % elimination of most of the test bacteria within 3 h and 6 h interaction. The partial characterization of the crude extracts by IR spectral analysis revealed possibility of terpenoid, long chain fatty acids and secondary amine derivatives compounds in the extracts. It is therefore recommended that further investigation should address the relationship between the structure of the active component of the extracts and the broad spectrum activity, as well as a rapid method for large scale production and purification and whether this group of antibiotics has any application in managing human infectious disease.

Stenotrophomonas maltophilia is increasingly emerging as an opportunistic pathogen of global concern. Due to its inherent resistance to several classes of antibiotics including carbapenems and its ability to acquire mobile resistance elements, treatment of infections caused by S. maltophilia is a constant challenge for clinicians. Trimethoprim-sulphamethoxazole (TMP-SMX) is the generally accepted antibiotic of choice for the treatment of infections caused by this organism, but resistance to the drug is increasingly being reported; hence, the need for alternative therapeutic options. In this study, the antimicrobial susceptibility profile of 110 commensal S. maltophilia isolates obtained from Nkonkobe municipality, Eastern Cape Province, Republic of South Africa was investigated. Twenty-one antibiotics including TMP-SMX and the newer fluoroquinolones; levofloxacin, gatifloxacin and moxifloxacin were included in the antibiotic panel. About 63.4 percent of the isolates were susceptible to TMP-SMX with a resistance rate of 28.2 percent. The fluoroquinolones were more effective with susceptibilities ranging from 76 percent to 94.7 percent. Resistance to the fluoroquinolones ranged from 1.3 percent to 2.7 percent. Levofloxacin was the most effective fluoroquinolone tested. Phenotypic dectection of extended spectrum β-lactamases (ESBLs) showed double disc synergy test (DDST) positivity in 59.5 percent of the isolates. Cefepime was the most sensitive indicator cephalosporin in the DDST with 77.3 percent of suspected ESBL-producing isolates showing cefepime-clavulanic acid synergy. Isolates exhibited nine different ESBL phenotypes, however, PCR amplification of the bla genes revealed four isolates that possessed genes belonging to the CTX-M group (CTX-M-1 and CTX-M-8 groups). ESBL genes are usually carried on mobile elements such as plasmids and transposons which may also bear genes that mediate resistance to aminoglycosides, tetracyclines, TMP-SMX and fluoroquinolones. ESBL positive isolates appeared more susceptible to the fluoroquinolones compared to TMP-SMX but there was no significant relationship between ESBL production and susceptibility to these drugs (p > 0.05). The newer fluoroquinolones are a possible alternative treatment option for S. maltophilia infections in this environment but further studies and clinical investigations are needed to determine the in vivo efficacy of these drugs.

No description available.

Susceptibility of 112 strains of Staphylococcus aureus obtained from Dameron Hospital, Stockton, California was tested with 18 antimicrobials . The MIC method was used with the following antimicrobials : tetracycline, oxacillin, penicillin, ampicillin, vancomycin, cefazolin, erythromycin, clindamycin, gentamycin, rifampin, trimethoprimsulfamethoxazole, chloramphenicol, and cefotaxime . The standard Kirby-Bauer disc diffusion method was used to test neomycin, tobramycin, and amikacin . Methicillin, oxacillin, and nafcillin were tested with a modified Kirby-Bauer method, which included the addition of a 4% salt supplement to the media, incubation at 32C, and readings at both 24 and 48 hours. Comparing results of this study with those of Hall (1975), suggested that resistance to the following antibiotics has increased: penicillin, ampicillin, erythromycin, neomycin, gentamycin, methicillin, oxacillin, nafcillin, cefazolin, and clindamycin . Resistance to tetracycline has decreased. No resistance to chloramphenicol or vancomycin was encountered in either study . Of the 112 strains studied, 13 . 4% were susceptible to all antibiotics tested. Twelve patterns of resistance were identified : 0 . 9% were resistant to neomycin only, 1.8% to erythromycin only, 63.9% to both penicillin and ampicillin, and 20 . 0% were multiply- resistant . Nine patterns of multiple-resistance were found, involving a minimum of three antibiotics and a maximum of nine . Three MRSA strains were identified from out-patient isolates; no in-patient isolates were methicillin-resistant . The study suggests that MRSA strains are not a problem at Dameron Hospital, but identification of this group would be more accurate if incubation of the MIC panels is maintained for at least 24 hours at ~35C . It was found that the MIC method of antimicrobial susceptibility testing is more reliable than the Kirby-Bauer disc diffusion method for detection of methicillin-resistance. Problems involved in identification of heteroresistant staphylococci are discussed .

Developing treatments for antibiotic resistant bacterial infections is among the highest priority public health challenges worldwide. Tetracyclines, one of the most important classes of antibiotics, have fallen prey to antibiotic resistance, necessitating the generation of new analogs. Many tetracycline analogs have been accessed through both total synthesis and semisynthesis, but key A- and C-ring tetracycline analogs remain inaccessible. New methods are needed to unlock access to these analogs, and heterologous biosynthesis in a tractable host such as Saccharomyces cerevisiae is a candidate method. C-ring analog biosynthesis can mimic nature’s biosynthesis of tetracyclines from anhydrotetracyclines, but challenges exist, including the absence of the unique cofactor F420 in common heterologous hosts. Chapter 1 provides background on antibiotics, and the tetracycline class in particular, and the metabolic engineering and directed evolution techniques available to us for heterologous expression of enzymes in yeasts. In Chapter 2, we describe the biosynthesis of tetracycline from anhydrotetracycline in S. cerevisiae heterologously expressing three enzymes from three bacterial hosts. Further, in Chapter 3, we highlight our Tang Laboratory collaborators’ work, where they reported the heterologous biosynthesis of a non-antibiotic fungal anhydrotetracycline derivative, TAN-1612, in S. cerevisiae from Aspergillus niger. We have built upon this system, allowing for the high-titer production of TAN-1612 in yeasts. Finally, in Chapter 4, we outline our efforts to convert TAN-1612 into a high titer tetracycline- and analog-producer by modifying the 2-, 4-, and 6-positions, proven critical for antibiotic activity. By hijacking biosynthetic hydroxylating and reducing enzymes, we attempted to modify the 6α-position, dearomatizing the C-ring. We also expressed heterologous enzymes within the TAN-1612 pathway that could furnish the 2-position with a carboxamido group instead of its natural hydrogen groups. By taking advantage of yeast’s natural biosynthetic pathways, we will create inexpensive, single-dose antibiotics, setting the stage to pursue yeast as a novel therapeutic. These state-of-the-art synthetic biology technologies will create entirely new paradigms, leading the charge against infections and diseases.

The spread of antibiotic-resistant microorganisms in the environment is recognized widely as an important public health issue, with concerns about future ability to treat infectious diseases. The main risk to public health is that the resistance genes are transferred from environmental bacteria to human pathogens. Safe water is one of the most important needs in public health in the twenty first century. The major health threat posed by drinking unsafe water is the transmission of infectious diseases, which are the leading causes of mortality and morbidity for children under the age of 5 and it is estimated to cause 1.5 million deaths annually in developing countries. In addition to the wide spread cases of water-borne diseases resulting from the contamination of water sources, concerns have been raised when these diseases fail to be cured due to development of resistance to most prescribed antibiotics by the contaminating microorganisms. It is now a well-established fact that E.coli is a significant cause of diarrheal illnesses both in infants and adults in many parts of the world. Data on clinical isolates is plenty while less attention has been given to environmental isolates of these enteric pathogens. Samples from the environment such as water may serve as probable reservoirs of these pathogens; this is compounded by the entry of functional compounds of antibiotics into waterways, through humans and animals that have ingested antibiotics. This is because antibiotics are not completely metabolized and may enter waterways through the waste products of these humans or animals.Studies on antimicrobial resistance is important in order to detect changes in patterns of resistance, implement control measures on the use of antimicrobial agents, and to prevent the spread of multidrug-resistant strains of bacteria. It also provides surveillance data for antibiotic resistances, necessary to define or update guidelines for empirical treatment, as well as a guide for appropriate antibiotic supplies. Study objectives: The objectives of this research were: (i) to determine the total and faecal coliform status of drinking water sources, as an indication of quality; (ii) to determine the bacteriological profile of bacteria flora in the drinking water sources; (iii) to determine prevalence and susceptibility profiles of antibiotic resistant water-borne E.coli; (iv) to investigate the virulence genes associated with multiple antibiotic resistant E. coli isolates; (v) to compare three laboratory based techniques: PCR, API 20E, and Culture based methods used for detection of E.coli and (vi) to determine the association between multiple antibiotic resistance and radiation sensitivity (D10). © University of South Africa 2014 VII Methodology: Four hundred and sixty four (464) water samples were collected for assessment between June 2011 and May 2012. The samples were collected from 57 sampling sites, from six different water sources including: boreholes (10), a canal (1), dams (15), hand-dug wells (15), a river (1), and streams (15). Total coliforms, faecal coliforms, and E. coli analysis were done by the MPN method. Bacteria isolation and identification were done using API 20E, conventional methods, and a PCR based DNA STRIP technology that allows simultaneous detection of virulence genes and confirmation of E. coli isolates. Antibiotic susceptibility testing was also conducted using the Kirby-bauer method. Radiation sensitivity was done using a cobalt 60 source. Results: The results obtained indicated that all the water sources were of poor quality in terms of microbial distributions with total coliform and faecal coliform counts ranging between 0 to 2.4x103 MPN/100ml. E. coli counts ranged between 10 to 7.9x101MPN/100ml. Disease risk assessment of the various water sources indicated that dam water sources presented a high disease risk, while borehole water sources had a low disease risk. A total of five hundred and twenty bacterial isolates (520) were obtained during the period of study. Three hundred and five (305) isolates representing 58.65% of the total were obtained during the dry season, as against (205) representing 41.35% in the rainy season. The most commonly occurring bacteria in the water samples was Klebsiella spp constituting 20%. The next most occurring organism was E. coli (18.7%). This was followed by Pseudomonas aeruginosa (15.61%), Enterobacter spp. (15.4%), Proteus vulgaris (13.1%), and Enterococcus faecalis (9.2%). The least isolated bacteria were Vibrio cholerae (1.2%) and Shigella spp. (1.2%). The prevalence of multi drug resistance E. coli was 49.48 %. E. coli isolates showed a high sistance patterns to the tested antibiotics. They were most resistant to penicillin (32.99%), cefuroxime (28.87%), erythromycin (23.71%), and tetracycline (21.45%). In contrast, they were susceptible to nitrofurantoin (93.8%), cefotaxime and amikacin (91.75%), gentamicin (90.7%), nalidixic acid (89.65%), ciprofloxacin (74.2%), chloramphenicol (69.07%), pipemidic acid (65.97%) and cefuroxime (52.58%). Sixty-three percent (63%) of the multidrug resistant E. coli strains recorded a multiple antibiotic resistance (MAR) index value of >0.2. Six (6%) percent of he multiple antibiotic resistant were eae virulence genes producing however, none of the E. coli isolates produced the stx1 and stx2 virulent gene. The analytical profile index (API) recorded specificity and sensitivity of 99.7% and 98.50 % respectively for the detection of E. coli. The © University of South Africa 2014 VIII culture/ biochemical based methods for detection of E coli recorded specificity of 81.82% and a sensitivity of 96.91%. There was no association (P> 0.05) between radiation sensitivity (D10) and antibiotic resistances. Conclusion: The study has confirmed that majority of the water sources used for drinking and domestic purposes in the study area are highly contaminated with high levels of faecal coliforms above the recommended standards. There were also resence of bacteria of public health importance in the water sources. Both animals and humans could be sources of faecal bacteria contamination of the drinking water sources. The study confirmed a high prevalence of multiple antibiotic resistances in E. coli isolates. The eae virulence gene was associated with some of the multiple resistant E. coli isolates. The study also concludes that API 20E has a high specificity and sensitivity close to that of the PCR. Lastly, There is no association between multiple antibiotic resistant indexes and radiation sensitivity (D10) of antibiotic resistant E. coli. / School of Environmental Sciences / D. Phil. (Environmental Science)

Thesis (PhD (Microbiology))--University of Stellenbosch, 2009. / Multidrug resistant strains of Staphylococcus aureus is presenting an increasing threat, especially immune compromised individuals. Many of these strains have developed resistance to newly approved drugs such as quinupristin-dalfopristin, linezolid and daptomycin. The search for alternative treatment, including bacteriocins (ribosomally synthesized antimicrobial peptides) of lactic acid bacteria is increasing . Lactococcus lactis subsp. lactis F10, isolated from freshwater catfish, produced a new nisin variant active against clinical strains of S. aureus. The operon encoding nisin F is located on a plasmid and the structural gene has been sequenced. The lantibiotic is closely related to nisin Z, except at position 30 where valine replaced isoleucine. The antimicrobial activity of nisin F against S. aureus was tested in the respiratory tract of Wistar rats. Non-immunosuppressed and immunosuppressed rats were intranasally infected with S. aureus K and then treated with either nisin F or sterile physiological saline. Nisin F protected immunosuppressed rats against S. aureus, as symptoms of an infection were only detected in the trachea and lungs of immunosuppressed rats treated with saline. The safety of intranasally administered nisin F was also evaluated and proved to have no adverse side effects. The potential of nisin F as an antimicrobial agent to treat subcutaneous skin infections was evaluated by infecting C57BL/6 mice with a bioluminescent strain of S. aureus (Xen 36). Immunosuppressed mice were treated with either nisin F or sterile physiological saline 24 h and 48 h after infection with subcutaneously injected S. aureus Xen 36. Histology and bioluminescence flux measurements revealed that nisin F was ineffective in the treatment of deep dermal staphylococcal infections. Non-infected and infected mice treated with nisin F had an influx of polymorphonuclear cells in the deep stroma of the skin tissue. This suggested that nisin F, when injected subcutaneously, may have modulated the immune system. Nisin F proved an effective antimicrobial agent against S. aureus-related infections in the respiratory tract, but not against subcutaneous infections. The outcome of nisin F treatment thus depends on the route of administration and site of infection.

Thesis (MSc (Biomedical Technology))--Cape Peninsula University of Technology, 2017. / Background: It is well established that Klebsiella pneumoniae (K. pneumoniae) is an opportunistic pathogenic organism that has been frequently identified as the cause of nosocomial and community acquired infections. Furthermore, studies have shown that over the last few decades strains of the genus Klebsiella have systematically developed resistance to numerous antibiotics. Aims and Methods: The primary aim of this study was to investigate the prevalence of K. pneumoniae in nosocomial and community isolates in the Western Cape province of South Africa. Various identification techniques such as the polymerase chain reaction (PCR) using the API 20 E, the VITEK®2 system, primers specific for the 16S-23S rDNA ITS region and the Matrix-assisted laser desorption/ionization Time of Flight Mass Spectrometry (MALDI-TOF MS) were compared for the identification of this pathogen. The VITEK 2 system was used to detect antibiotic resistant profiles of the K. pneumoniae isolates and to identify the extended spectrum beta-lactamase (ESBL) phenotypic among these isolates. The PCR was used to detect Beta-lactam genes viz. CTX-M (blaCTX-M), TEM (blaTEM) and SHV (blaSHV) respectively in both the genome and plasmid DNA of K. pneumoniae using gene specific primers. Results: In total 57 agar plate bacterial cultures or glycerol stock bacterial cultures were obtained during 2011. Of the 57 isolates, the API 20 E test identified 47 (82.5%) of the isolates (n = 57) as K. pneumoniae while 10 isolates (17.5%) were identified as Raoultella species. The VITEK 2 method and PCR identified all 57 isolates as K. pneumoniae (100%). Of the isolates, 82.5% (47/57) were positively identified as Klebsiella species, 14% (8/57) were identified as Klebsiella variicola and 3.5% (2/57) were shown as no reliable identification (NRI) when using the MALDI-TOF MS. Examination of the 57 isolates using primers specific for the CTX-M (blaCTX-M), TEM (blaTEM) and SHV (blaSHV) respectively showed the following: PCR amplicons for the TEM gene were produced successfully for 46 (81%) of the 57 isolates included in this project, while 11 (19%) of the samples did not yield any TEM amplicons; PCR amplicons for the blaSHV gene were obtained successfully for 56 (98%) of the 57 DNA samples, while 1 sample (2%) did not yield any SHV amplicons; and PCR amplicons for the blaCTX-M gene were produced successfully by 89% (n = 51) of the DNA samples included in this project, while 11% (n = 6) did not yield any CTX-M amplicon. Extended-spectrum beta-lactamase phenotypes had been confirmed in 84% (n = 48) K. pneumoniae isolates while nine isolates were found to be non-ESBL. Resistance rates for these 48 isolates were high and showed resistance patterns of: Amoxicillin/Ampicillin, Amoxycillin/Clavulanate, Ceftriaxone/Cefotaxime, Cefuroxime/Cefprozil and Ceftazidime (100%, n = 48); Piperacillin/Tazobactam and Cefoxitin (98%, 47/48); Cefepime (96%, 46/48); Aztreonam (94%; 45/48); Tobramycin (81%, 39/48); Gentamycin and Ciprofloxacin (77%, 37/48); Trimethoprim/Sulfamethoxazole (67%, 32/48); and Tigecycline (25% 12/48). Conclusion: For the analysis by all four methods employed, a total agreement of 68.4% was obtained, indicating the positive identification of K. pneumoniae in 39 of the 57 samples analysed. An average agreement of 28.1% was then obtained for the comparison of results generated for three of the methods utilised, while a 3.5% average agreement was obtained for at least two methods. Furthermore, all four methods agreed that 82.5% of the isolates were Klebsiella species while three methods agreed that 17.5% of the isolates were Klebsiella species. Based on the results obtained in the current study, PCR and VITEK 2 were the methods of choice for the identification of K. pneumoniae. The current study also showed, that ESBL-K. pneumoniae strains are present in the Western Cape province, South Africa; with high resistance profiles to numerous antibiotics including the Cephalosporins.

Abstract not available